The application of nanomaterials as carrier systems to deliver imaging reagents and/or drugs has gained momentum in the medical field. Nanoparticles are advantageous because their large surface-area-to-volume ratio allows functionalization with multiple different payloads and ligands. Nanoparticles are used to partition cargos between diseased and healthy tissue, ideally avoiding healthy tissues or at least minimizing the accumulation of toxic substances in healthy organs. Disease targeting (e.g., to cancer, inflammation, or infection), is achieved making use of the unique biological features that distinguish the microenvironment of diseased cells from healthy cells. For example, based on their size, nanoparticles home to solid tumors as a result of their leaky tumor blood vessels and the associated enhanced permeability and retention effects. Perrault et al., Nano Letters 9, 1909-1915 (2009). Other targeting strategies include the use of receptor-specific ligands to direct the nanocarrier to receptors selectively over-expressed at the target disease site. Ruoslahti E, Biochem Soc Trans, 32(Pt3), 397-402 (2004).
When it comes to cargo-loading and cargo-release, many different chemistries and mechanisms have been developed that control loading efficiency, affinity, and release rates; the choice of chemistry typically depends on the disease profile, cargo molecule, and carrier system of choice. Many different carrier systems are currently under investigation and development for drug delivery and tissue-specific imaging; each system has its advantages and disadvantages with regard to physiochemical properties, biodistribution and clearance, pharmacokinetic behavior, immunogenicity, and toxicity.
The inventors have focused on the development of bionanoparticles derived from plant viruses, also termed viral nanoparticles (VNPs). There are many novel types of VNPs under development, with those based on bacteriophages and plant viruses favored because they are considered safer in humans than mammalian viruses. Manchester M, Singh P, Adv Drug Deliv Rev 58(14), 1505-1522 (2006). Preclinical studies in mice have shown that plant viruses can be administered at doses of up to 100 mg (1016 VNPs) per kg body weight without signs of toxicity. Singh et al., J Control Release, 120, 41-50 (2007). Like other protein-based nanomaterials they are immunogenic. However, strategies such as PEGylation can be used to overcome the immunogenicity of VNPs. Raja et al., Biomacromolecules, 3, 472-476 (2003). VNPs are genetically encoded and self-assemble into discrete and monodisperse structures with a precise shape and size. Many virus structures are understood at atomic resolution, allowing the development of protocols for high-precision VNP tailoring. This level of quality control cannot yet be achieved with synthetic nanoparticles. VNPs can be modified with targeting ligands and/or cargos using at least five approaches: genetic engineering, bioconjugate chemistry, self-assembly, mineralization, and infusion techniques. Pokorski J K, Steinmetz N F, Mol Pharm, 8, 29-43 (2011).
Cowpea mosaic virus (CPMV) is a plant picornavirus typically produced in black-eyed pea plants. CPMV capsids measure 30 nm in diameter and are comprised by 60 copies each of a small (S) and large (L) protein encapsulating a bipartite, single stranded, positive-sense RNA genome. CPMV has been extensively studied, developed, and tested for applications in the medical field. Bioconjugate chemistries on CPMV's exterior and interior surfaces are well established and its in vitro and in vivo properties are well understood. Wu et al., Nanoscale, 4, 3567-3576 (2012). CPMV naturally is taken up by mammalian cells through interactions with surface-expressed vimentin. Koudelka et al., PLoS Pathog, 5, e1000417 (2009). This unique property can be used to target CPMV to endothelial cells for vascular imaging and tumor vessel mapping (Lewis et al., Nature Medicine, 12, 354-360 (2006)), targeting vimentin-expressing cancer cells in vitro or in vivo (Steinmetz et al., Nanomedicine (Lond), 6, 351-364 (2011)), as well as targeting and imaging sites of inflammation, such as atherosclerotic plaques or infections of the central nervous system. Re-targeting of CPMV to receptors of interest can also be achieved through tailoring the surface chemistry with appropriate targeting ligands. Hovlid et al., Nanoscale 4, 3698-3705 (2012).
More recently, the application of CPMV as a carrier for drug delivery has been demonstrated. Cytotoxicity of CPMV nanoparticles chemically modified with multiple copies of the chemotherapeutic drug doxorubicin has been investigated. Aljabali et al., Mol. Pharm., 10, 3-10 (2013). However, multistep chemical modification procedures can be cumbersome, low yielding, and costly. Accordingly, there is a need for methods of using CPMV nanoparticles for drug delivery without requiring the use of multistep chemical modification procedures.